OZONATION OF DNA FORMS ADDUCTS - A P-32 DNA LABELING AND THIN-LAYER CHROMATOGRAPHY TECHNIQUE TO MEASURE DNA ENVIRONMENTAL BIOMARKERS

Little direct documented evidence of ozone's genotoxicity exists. Deoxyribonucleic acid (DNA)adducts are produced by environmental toxic agents, including ozone. We have described a modified thin-layer chromatography (TLC) technique that can assess adduct formation as a biomarker of ozone injury. This requires P-32-labeling DNA, digestion of deoxynucleotides (dNMPs), and separation in two-dimensional PEI-cellulose TLC. We have applied this technique to control DNAs, to control DNA in solution exposed to acute ambient ozone, and to control DNA exposed to acute bubbled-through ozone (2 ppm for 24 h). We detected stable DNA adducts, including hydroxymethyluracil (HMU), thymine glycol (TG), 8-hydroxyguanine (8-OHG), and demonstrated, as yet, unidentified adducts that may serve as a ''fingerprint'' pattern of DNA adduction. This technique quantifies low-molecular-mass DNA adducts, both in vivo and in vitro, with potential applications to environmental toxicology.